AB0061 PHARMACOKINETICS OF METHOTREXATE POLYGLUTAMATES IN PERIPHERAL BLOOD MONONUCLEAR CELLS OF RA PATIENTS IS SIMILAR AFTER SUBCUTANEOUS OR ORAL ADMINISTRATION IN THE METHOTREXATE MONITORING (MeMo) TRIAL
Authors: R. Hebing, M. Lin, E. Struys, S. Mahmoud, I. Muller, W. Lems, B. van den Bemt, G. Jansen, R. De Jonge, M. Nurmohamed
Journal: Annals of the Rheumatic Diseases (2022)

Background Pharmacokinetics of methotrexate (MTX) after oral and subcutaneous administration to RA patients differs: MTX levels in plasma and MTX-polyglutamate (MTX-PG) accumulation in erythrocytes are higher during equidosed subcutaneous compared to oral MTX treatment. (1,2) No data are available whether administration route of MTX differentially impacts the intracellular concentrations of MTX-PGs in peripheral blood mononuclear cells (PBMCs) during MTX therapy. Objectives To investigate the pharmacokinetics of MTX-PGs in PBMCs of newly diagnosed RA patients receiving oral or subcutaneous MTX in the early phase (1, 2, 3 and 6 months) of MTX treatment. Methods In a clinical prospective cohort study (MeMo study (NTR7149)), RA patients were administered oral (n=24) or subcutaneous (n=22) MTX up to 25 mg MTX/week, as described before. (1) At 1, 2, 3 and 6 months after the start of therapy, PBMCs were isolated via Ficoll density gradient centrifugation. Individual MTX-PG forms (MTX-PG 1-6 ) in PBMCs were analyzed by a UPLC-MS/MS method including custom-made stable isotopes of MTX-PG 1-6 as internal standards (3). UPLC-MS/MS measurements of the PBMCs were performed with a Waters Acquity BEH C18 column coupled to an AB Sciex 6500+ with the ESI operating on the positive mode. Dosing, concomitant treatments and DAS28-ESR assessments were in conformity with clinical practice. (4) Results 46 consecutive patients were included in this study; 76% female, mean age: 57.8 years, BMI: 25.8, smokers: 20%, mean baseline DAS28-ESR: 3.5, as described before. (1) MTX dose at baseline was 10.5 mg (SD: 1.5) for both groups, 15.4 mg (4.4) and 16.8 mg (1.8) at 1 month, 22.8 mg (3.9) and 22.4 mg (5.2) at 2 months, 20.1 mg (6.3) and 20.8 mg (5.6) at 3 months, and 19.7 mg (6.1) and 18.5 mg (6.7) at 6 months for oral and subcutaneous use, respectively. MTX-PG analyses in PBMCs for individual and total MTX-PGs revealed no significant differences between oral and subcutaneous administration groups at 1, 2, 3, and 6 months (Figure 1). Linear regression of LN transformed MTX-PG levels in PBMCs and administration route, corrected for age, baseline DAS28, smoking, BMI, eGFR and MTX dose, showed a trend towards higher MTX-PG levels in PBMCs after subcutaneous MTX administration compared to oral administration (data not shown). MTX-PG distribution in PBMCs was mainly composed of MTX-PG 1 (58%), and to a lesser extent MTX-PG 2 (27%) and MTX-PG 3 (15%). Longer chain MTX-PGs beyond MTX-PG 4 were detectable in PBMCs, but at levels lower than MTX-PG 1-3 (mean: 4.0 – 6.7 fmol/10 6 cells). Total MTX-PG accumulation in PBMCs was approximately 10-20 fold higher than in erythrocytes. PBMC accumulation was rather stable, whereas RBC MTX-PG accumulation increased between 1 to 3 months to reach a plateau (Figure 1). Figure 1. Loess regression of MTX-PG concentrations in PBMCs (MTX-PG1-3) and RBCs (MTX-PG1-6) of RA patients during the first 6 months of oral or subcutaneous MTX administration. At 6 months, 18 patients using oral and 18 patients using subcutaneous MTX were still continuing MTX treatment. Means (lines) and SE (grey areas) are depicted. Conclusion This study demonstrated that MTX-PG accumulation in PBMCs early on in the MTX treatment of RA patients was not significantly different between oral or subcutaneous MTX administration routes. References [1]RCF Hebing et al, Arthritis Rheum (2021); 60:339-348 [2]M Hoekstra et al, J Rheumatol (2004); 31:645-8 [3]E Den Boer et al, Anal Bioanal Chem (2013); 405: 1673-1681 [4]J Smolen et al, Ann Rheum Dis (2020); 79:685-699 Acknowledgements We would like to thank all participating patients and Pfizer (grant 53233663 / WI230458), NVKC (Noyons grant) and AmsterdamUMC (AI&amp;II extension grant). Disclosure of Interests Renske Hebing Grant/research support from: Pfizer, grant number 53233663 / WI230458, Amsterdam UMC (AI&amp;II extension grant), NVKC (Netherlands Society for Clinical Chemistry, Noyons grant), Marry Lin: None declared, Eduard Struys: None declared, Sohaila Mahmoud: None declared, Ittai Muller: None declared, WIllem Lems: None declared, Bart van den Bemt Speakers bureau: Pfizer, UCB, Sanofi-Aventis, Galapagos, Amgen and Eli Lilly, Gerrit Jansen: None declared, Robert De Jonge Grant/research support from: NVKC (Netherlands Society for Clinical Chemistry, Noyons grant), Michael Nurmohamed Grant/research support from: Pfizer grant